Wound dressings and applications thereof

ABSTRACT

In one aspect, compositions and wound dressings are described herein. In some embodiments, a composition or wound dressing described herein comprises a mesh formed from a plurality of biodegradable polymer fibers; a first active agent dispersed in the biodegradable polymer fibers; a plurality of biodegradable polymer particles disposed in the mesh; and a second active agent dispersed in the biodegradable polymer particles. The particles can be disposed within the interiors of the fibers of the mesh or between the fibers of the mesh. In another aspect, a composition or wound dressing described herein comprises a first perforated mesh formed from a first plurality of biodegradable polymer fibers; and a second perforated mesh formed from a second plurality of biodegradable polymer fibers, wherein the second perforated mesh is disposed on the first perforated mesh in a stacked configuration and the first and second perforated meshes have different degrees of perforation.

CROSS REFERENCE TO RELATED APPLICATION

This application claims priority pursuant to 35 U.S.C. § 119 to U.S.Provisional Patent Application Ser. No. 61/826,328, filed on May 22,2013, which is hereby incorporated by reference in its entirety.

STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH

This invention was made with government support under contract EB012575awarded by the National Institute of Biomedical Imaging andBioengineering (NIBIB), and contract DMR1313553 awarded by the NationalScience Foundation (NSF). The government has certain rights in theinvention.

FIELD

This invention relates to compositions and wound dressings and, inparticular, to compositions and wound dressings comprising biodegradablepolymer fibers and biodegradable polymer particles.

BACKGROUND

Wound healing is a dynamic and complex process involving extracellularmatrix (ECM), cytokines, blood cells, and other biological species.Further, wound healing can include three overlapping phases:inflammation, tissue regeneration, and tissue remodeling. Due to thepathological and physiological complexity of the wound healing process,perfect tissue regeneration can be difficult to achieve, especially forskin wounds and chronic wounds such as diabetic ulcers and hernias.Hernia repair is one of the most common surgeries in the United States,where up to 27% of men and 3% women are affected by hernias.Unfortunately, some existing hernia repair procedures and devices, suchas non-biodegradable hernia repair meshes, can themselves cause chronicpain and/or recurrence of the hernia. For example, some existing meshesexhibit poor cell infiltration into the mesh and/or poor mechanicalperformance.

Further, many existing wound dressings act only as temporary barriersfor hemostasis to protect the patient from infection and do nototherwise promote wound healing. In addition, some natural and syntheticskin graft applications can be expensive, require extensivepost-procedure care, and/or fail to provide full skin functionality.Therefore, there exists a need for improved wound dressings and methodsof treating wounds such as skin wounds, diabetic ulcers, and hernias.

SUMMARY

In one aspect, compositions and wound dressings are described hereinwhich, in some embodiments, can provide one or more advantages comparedto some other compositions and wound dressings. For example, in someembodiments, a composition or wound dressing described herein canpromote more rapid wound healing through the controlled release of aplurality of active agents according to a desired release profile,including a bimodal or partially overlapping release profile. Acomposition or wound dressing described herein can also promote woundhealing by presenting a compositional gradient to a wound site, such asa porosity or perforation gradient. Moreover, in some cases, acomposition or wound dressing described herein can provide a fibrousscaffold for supporting cell growth, including regenerated cell growth.Such a fibrous scaffold, in some instances, can mimic the extracellularmatrix (ECM) of living organisms. In addition, a fibrous scaffoldprovided by a composition or wound dressing described herein can alsoexhibit antimicrobial properties. Moreover, compositions or wounddressings described herein can provide one or more of the foregoingadvantages simultaneously. For example, in some cases, a composition orwound dressing described herein can serve as a scaffold to support woundhealing while also releasing multiple active agents at various phases ofwound healing. In some embodiments, a composition or wound dressingdescribed herein can simultaneously provide biochemical stimulation,cell growth support, and bacteria inhibition.

In some embodiments, a composition described herein comprises abiodegradable polymer fiber, a first active agent dispersed within thefiber, a plurality of biodegradable polymer particles dispersed withinthe fiber, and a second active agent dispersed within the polymerparticles. In addition, in some cases, a composition or wound dressingdescribed herein comprises a mesh formed from a plurality ofbiodegradable polymer fibers; a first active agent dispersed in thebiodegradable polymer fibers; a plurality of biodegradable polymerparticles disposed in the mesh; and a second active agent dispersed inthe biodegradable polymer particles. In some embodiments, the particlesare disposed within the interiors of the fibers of the mesh.Alternatively, in other cases, the particles are disposed between thefibers of the mesh. Additionally, the mesh of a composition or wounddressing described herein can be a non-woven mesh.

As described further hereinbelow, a composition or wound dressing havinga structure described herein, in some embodiments, can provide abifurcated, bimodal, or temporally separated delivery of the first andsecond active agents to a wound or other biological compartment when thewound dressing is disposed on the wound or in the biologicalcompartment.

One or more active agents of a composition or wound dressing describedherein, in some cases, can comprise a growth factor, such as a growthfactor for angiogenesis, wound healing, or bone growth. Moreover, insome embodiments, the identity of one or more growth factors is selectedto achieve one or more desired biological effects, including in adesired temporal sequence. In some instances, for example, a firstactive agent is selected to achieve a first biological effect, such aspromotion of blood vessel growth, and a second active agent is selectedto achieve a second biological effect, such as wound healing or thepromotion of bone growth, that may desirably be temporally separatedfrom the first biological effect.

Additionally, in some embodiments, the biodegradable polymer fibers of acomposition or wound dressing described herein comprise one or moreantimicrobial polymer fibers. In some cases, the biodegradable polymerfibers comprise one or more of chitosan, carboxymethyl chitosan (CMC),poly(ethylene oxide), and collagen. In other embodiments, thebiodegradable polymer fibers of a composition or wound dressingdescribed herein comprise one or more polymers comprising a citratemoiety. Moreover, in some instances, the biodegradable polymer fibersare nanofibers having an average diameter between about 50 nm and about1000 nm. Similarly, in some cases, the biodegradable polymer particlesof a composition or wound dressing described herein are nanoparticleshaving an average size between about 10 nm and about 200 nm.

In another aspect, compositions or wound dressings described hereincomprise a stack of biodegradable polymer fiber meshes that may or maynot comprise biodegradable polymer particles and/or active agents. Insome embodiments, such a stack of meshes is arranged to provide aproperty gradient in the z-direction, as described further hereinbelow.For example, in some cases, a composition or wound dressing describedherein comprises a first perforated mesh formed from a first pluralityof biodegradable polymer fibers; and a second perforated mesh formedfrom a second plurality of biodegradable polymer fibers, wherein thesecond perforated mesh is disposed on the first perforated mesh in astacked configuration and the first perforated mesh has a higher degreeof perforation than the second perforated mesh. Additionally, in someembodiments, such a composition or wound dressing further comprises athird perforated mesh formed from a third plurality of biodegradablepolymer fibers, wherein the third perforated mesh is disposed on thesecond perforated mesh in a stacked configuration and the thirdperforated mesh has a higher degree of perforation than the firstperforated mesh and the second perforated mesh. Moreover, as describedfurther hereinbelow, compositions or wound dressings described hereincan further comprise additional perforated meshes or non-perforatedmeshes in a stacked configuration. For example, in some cases, acomposition or wound dressing described herein further comprises afourth mesh formed from a fourth plurality of biodegradable polymerfibers, wherein the fourth mesh is non-perforated or has a lower degreeof perforation than the third perforated mesh. A wound dressing havingsuch a structure, in some embodiments, can provide a physical barrier tocomplete tissue penetration of the wound dressing on the side of thewound dressing farther from the wound.

Additionally, if desired, one or more meshes of a stack described hereincan have a structure described hereinabove for wound dressingscomprising active agents. For example, in some cases, an active agent isdispersed in the biodegradable polymer fibers of the first perforatedmesh and/or the second perforated mesh. In some instances, a pluralityof biodegradable polymer particles is disposed in the first perforatedmesh and/or the second perforated mesh. Moreover, in some cases, asecond active agent is dispersed in the biodegradable polymer particles.

In another aspect, methods of treating a wound are described herein. Insome embodiments, a method of treating a wound described hereincomprises applying a composition or wound dressing described hereinaboveto a surface of the wound, which may be a skin wound, diabetic ulcer, orhernia. For example, in some embodiments, the wound dressing comprises amesh formed from a plurality of biodegradable polymer fibers; a firstactive agent dispersed in the biodegradable polymer fibers; a pluralityof biodegradable polymer particles disposed in the mesh; and a secondactive agent dispersed in the biodegradable polymer particles. Such amethod, in some cases, can further comprise at least partially degradingthe biodegradable polymer fibers to release one or more active agentsinto the wound.

These and other embodiments are described in more detail in the detaileddescription which follows.

BRIEF DESCRIPTION OF THE FIGURES

FIG. 1 illustrates schematically a wound dressing and a method oftreating a wound according to one embodiment described herein.

FIGS. 2A and 2B illustrate scanning electron microscope (SEM) images ofmeshes of wound dressings according to some embodiments describedherein.

FIG. 2C illustrates a fluorescence image of a wound dressing accordingto one embodiment described herein.

FIG. 2D illustrates plots of the diameters of polymer fibers of wounddressings according to some embodiments described herein.

FIG. 3 illustrates plots of the release profiles of active agents ofwound dressings according to some embodiments described herein.

FIG. 4 illustrates plots of cell proliferation following treatment of awound according to some embodiments described herein.

FIGS. 5A and 5B illustrate plots of the antimicrobial properties ofwound dressings according to some embodiments described herein.

FIG. 6A illustrates photographs of wounds treated according to someembodiments of methods described herein.

FIG. 6B illustrates plots of wound healing as a function of timeaccording to some embodiments of methods described herein.

FIG. 7A illustrates staining images of wounds treated according to someembodiments of methods described herein.

FIGS. 7B-7D illustrate plots of the results of treating a woundaccording to some embodiments of methods described herein.

FIG. 8A illustrates staining images of wounds treated according to someembodiments of methods described herein.

FIGS. 8B and 8C illustrate plots of the results of treating a woundaccording to some embodiments described of methods herein.

FIG. 9 illustrates schematically a method of making a wound dressingaccording to one embodiment described herein.

FIGS. 10A-D illustrate SEM images of perforated meshes of wounddressings according to some embodiments described herein.

FIGS. 11A-D illustrate plots of mechanical properties of meshes of wounddressings according to some embodiments described herein.

DETAILED DESCRIPTION

Embodiments described herein can be understood more readily by referenceto the following detailed description, examples, and figures. Elements,apparatus, and methods described herein, however, are not limited to thespecific embodiments presented in the detailed description, examples,and figures. It should be recognized that these embodiments are merelyillustrative of the principles of the present invention. Numerousmodifications and adaptations will be readily apparent to those of skillin the art without departing from the spirit and scope of the invention.

In addition, all ranges disclosed herein are to be understood toencompass any and all subranges subsumed therein. For example, a statedrange of “1.0 to 10.0” should be considered to include any and allsubranges beginning with a minimum value of 1.0 or more and ending witha maximum value of 10.0 or less, e.g., 1.0 to 5.3, or 4.7 to 10.0, or3.6 to 7.9.

All ranges disclosed herein are also to be considered to include the endpoints of the range, unless expressly stated otherwise. For example, arange of “between 5 and 10” should generally be considered to includethe end points 5 and 10.

Further, when the phrase “up to” is used in connection with an amount orquantity, it is to be understood that the amount is at least adetectable amount or quantity. For example, a material present in anamount “up to” a specified amount can be present from a detectableamount and up to and including the specified amount.

I. Compositions and Wound Dressings

In one aspect, compositions and wound dressings are described herein. Insome embodiments, a composition described herein comprises abiodegradable polymer fiber, a first active agent dispersed within thefiber, a plurality of biodegradable polymer particles dispersed withinthe fiber, and a second active agent dispersed within the polymerparticles. In addition, in some cases, a composition or wound dressingdescribed herein comprises a mesh formed from a plurality ofbiodegradable polymer fibers; a first active agent dispersed in thebiodegradable polymer fibers; a plurality of biodegradable polymerparticles disposed in the mesh; and a second active agent dispersed inthe biodegradable polymer particles. In some cases, the particles aredisposed within the fibers of the mesh, such that the fibers completelyor substantially completely contain the particles within the interiorvolumes of the fibers. In such instances, the polymer particles can bepresent within the polymer fibers in any amount not inconsistent withthe objectives of the present disclosure. In some embodiments, forexample, the biodegradable polymer particles are present within thefibers in an amount up to about 30 weight percent, based on the totalweight of the fibers plus the particles. In other cases, thebiodegradable polymer particles are present within the fibers in anamount up to about 25 weight percent, up to about 20 weight percent, upto about 15 weight percent, up to about 10 weight percent, or up toabout 5 weight percent, based on the total weight of the fibers plus theparticles. In some embodiments, the biodegradable polymer particles arepresent within the fibers in an amount between about 1 weight percentand about 30 weight percent, between about 5 weight percent and about 25weight percent, between about 5 weight percent and about 20 weightpercent, between about 5 weight percent and about 15 weight percent, orbetween about 10 weight percent and about 20 weight percent, based onthe total weight of the fibers plus the particles. Fibers comprisingsuch amounts of particles, in some embodiments, can be smooth, uniform,and substantially beadless composite fibers.

In some cases, the biodegradable polymer particles of a wound dressingdescribed herein are disposed between the fibers of the mesh. Particlesthat are disposed between the fibers of the mesh can be physicallyentrapped in the mesh and/or chemically bonded to the outer surface ofthe fibers of the mesh, as opposed to being incorporated into theinterior volume of the fibers. In such cases, the biodegradable polymerparticles can be present in the mesh in any amount not inconsistent withthe objectives of the present disclosure. In some embodiments, thebiodegradable polymer particles can be present in the mesh in an amountup to about 80 weight percent, up to about 70 weight percent, up toabout 50 weight percent, up to about 40 weight percent, up to about 30weight percent, up to about 20 weight percent, up to about 10 weightpercent, or up to about 5 weight percent, based on the total weight ofthe mesh plus the particles. In some cases, the biodegradable polymerparticles are present in the mesh in an amount between about 1 weightpercent and about 80 weight percent, between about 5 weight percent andabout 70 weight percent, between about 10 weight percent and about 50weight percent, between about 10 weight percent and about 40 weightpercent, or between about 10 weight percent and about 20 weight percent,based on the total weight of the mesh plus the particles.

As described further hereinbelow, a wound dressing having a structuredescribed herein, in some embodiments, can provide a bifurcated,bimodal, or temporally separated delivery of the first and second activeagents to a wound or other biological compartment when the wounddressing is disposed in contact with the wound or other biologicalcompartment. In some cases, such a release profile can be achieved evenwhen the first and second active agents are chemically similar and/orwhen the biodegradable polymer fibers are chemically similar to thebiodegradable polymer particles. For example, first and second activeagents having the same or similar hydrophobicity, hydrophilicity,electrostatic charge, and/or hydrodynamic size can nevertheless exhibitdifferent in vivo or in vitro release profiles when included in a wounddressing having a structure described herein. An “in vivo or in vitrorelease profile,” for reference purposes herein, describes the amount orconcentration of an active agent that is released from a wound dressingover time (t) when the wound dressing is disposed in an in vivo or invitro environment, respectively. Such release of an active agent mayoccur due to diffusion of the active agent out of the biodegradablepolymer fibers or particles in which the active agent is dispersed.Release of an active agent may also occur due to degradation of thebiodegradable polymer fibers or particles.

In some embodiments, the in vivo or in vitro release profile of thefirst active agent of a wound dressing described herein differs from thein vivo or in vitro release profile of the second active agent. Forexample, in some cases, the in vivo or in vitro release profile of thefirst active agent and the in vivo or in vitro release profile of thesecond active agent overlap by less than about 70%. In some embodiments,the release profiles overlap by less than about 50%, less than about40%, less than about 30%, less than about 25%, less than about 20%, lessthan about 15%, less than about 10%, less than about 5%, or less thanabout 1%. In some instances, the release profiles are entirelynon-overlapping.

The percent “overlap” of active agent release profiles can be based onthe total area of the in vivo or in vitro release profile curves of theactive agents, as described further hereinbelow. For example, a firstactive agent may be completely released from a wound dressing describedherein beginning at 3 days after placement of the wound dressing andending at 7 days after placement. A second active agent may begin to bereleased from the wound dressing after 7 days after placement. In suchan instance, the in vivo or in vitro release profile of the first activeagent would overlap the in vivo or in vitro release profile of thesecond active agent by 0%. Alternatively, if half of the total amount ofthe first active agent were released between t=6 days and t=7 days (withthe other half being released prior to t=6 days), and half of the totalamount of the second active agent were released between t=6 days and t=7days (with the other half being released after t=7 days), then the invivo or in vitro release profiles of the first and second active agentswould overlap by 50%.

In general, a desired overlap between the in vivo or in vitro releaseprofiles of active agents described herein can be selected based on oneor more of the following: the chemical composition of each of the activeagents, the chemical composition of the biodegradable polymer fibers,the chemical composition of the biodegradable polymer particles, theamount of active agent dispersed in each of the biodegradable polymerfibers and particles, the physical dimensions of the biodegradablepolymer fibers and particles, and the biodegradation rates of thebiodegradable polymer fibers and particles. Further, in some cases, eachof the foregoing features can be used independently to increase ordecrease the overlap of active agent release profiles. For example, toachieve a higher percent overlap, the chemical composition of the firstand second active agents can be selected to exhibit the same or similarhydrophobicity, hydrophilicity, electrostatic charge, and/orhydrodynamic size. A higher percent overlap can also be achieved byproviding biodegradable polymer fibers and biodegradable polymerparticles having similar chemical compositions, similar biodegradationrates, and/or similar sizes. In contrast, to achieve a lower percentoverlap of in vivo or in vitro release profiles, the foregoingproperties of the first and second active agents can differ. It is alsopossible to achieve a lower percent overlap of in vivo or in vitrorelease profiles by dispersing a hydrophobic first active agent inhydrophilic biodegradable polymer fibers and dispersing a hydrophilicsecond active agent in hydrophilic biodegradable polymer particles. Adesired overlap of release profiles can be achieved in other ways aswell.

Similarly, the absolute release rates of the first and second activeagents can also be independently selected based on one or more of theforegoing factors, including one or more of the chemical composition ofeach of the active agents, the chemical composition of the biodegradablepolymer fibers, the chemical composition of the biodegradable polymerparticles, the amount of active agent dispersed in each of thebiodegradable polymer fibers and particles, the physical dimensions ofthe biodegradable polymer fibers and particles, and the biodegradationrates of the biodegradable polymer fibers and particles. In some cases,the first active agent of a wound dressing described herein has arelease half-life of less than about 4 days, less than about 3 days,less than about 2 days, or less than about 1 day. In some instances, thefirst active agent of a wound dressing described herein has a releasehalf-life between about 0.5 days and about 5 days, between about 0.5days and about 4 days, between about 1 day and about 4 days, or betweenabout 1 day and about 3 days. In some embodiments, the first activeagent has a release half-life of less than 1 day. Additionally, in somecases, the second active agent has a release half-life longer than therelease half-life of the first active agent. For example, in someembodiments, the second active agent of a wound dressing describedherein has a release half-life of greater than about 2 days, greaterthan about 3 days, greater than about 4 days, greater than about 5 days,greater than about 6 days, greater than about 7 days, or greater thanabout 10 days. In some instances, the second active agent has a releasehalf-life between about 1 day and about 10 days, between about 2 daysand about 9 days, between about 3 days and about 8 days, between about 3days and about 7 days, between about 4 days and about 10 days, betweenabout 4 days and about 7 days, or between about 4 days and about 6 days.Thus, in some embodiments, a wound dressing described herein can providea rapid release of a first active agent (such as when the releasehalf-life of the first active agent is less than about 2 days or lessthan about 1 day), followed by a slower, sustained release of a secondactive agent (such as when the release half-life of the second activeagent is greater than about 5 days). Moreover, in some cases, suchrelease rates can be obtained even when the first and second activeagents have the same or similar hydrophobicity, hydrophilicity,hydrodynamic size, and/or electrostatic charge, including relative tothe biodegradable polymer fibers and particles in which the activeagents are dispersed. The “release half-life” of an active agent, forreference purposes herein, refers to the amount of time needed for halfthe total amount of the active agent to be released from the wounddressing following placement of the wound dressing in an in vivo or invitro environment.

Further, in some embodiments, the first active agent of a wound dressingdescribed herein has a release profile wherein at least about 30%, atleast about 50%, or at least about 60% by weight of the active agent isreleased within 30 minutes of disposing the wound dressing in an in vivoor in vitro environment. In some cases, the first active agent has arelease profile wherein between about 30% and about 70% or between about30% and about 65% of the active agent is released within 30 minutes ofdisposing the wound dressing in an in vivo or in vitro environment. Insome embodiments, the first active agent has a release profile whereinat least about 90%, at least about 95%, or at least about 99.9% byweight of the active agent is released within 3 days of disposing thewound dressing in an in vivo or in vitro environment.

Similarly, in some cases, the second active agent of a wound dressingdescribed herein has a release profile wherein less than about 30%, lessthan about 20%, or less than about 15% by weight of the active agent isreleased during the first 24 hours after disposing the wound dressing inan in vivo or in vitro environment. In some cases the second activeagent has a release profile wherein between about 1% and about 30% orbetween about 5% and about 25% of the active agent is released duringthe first 24 hours after disposing the wound dressing in an in vivo orin vitro environment. Additionally, in some embodiments, the secondactive agent has a sustained release profile in vivo or in vitro.

Further, although wound dressings having two active agents are describedherein, it is to be understood that wound dressings can also comprisemore than two active agents. For example, in some embodiments, aplurality of differing first active agents is disposed in the polymerfibers of a wound dressing, and/or a plurality of differing secondactive agents is disposed in the polymer particles of a wound dressingdescribed herein.

Turning now to specific components of some wound dressings, wounddressings described herein, in some embodiments, comprise a mesh formedfrom biodegradable polymer fibers. Any biodegradable polymer fibers notinconsistent with the objectives of the present disclosure may be used.A biodegradable polymer, in some embodiments, degrades in vivo tonon-toxic components which can be cleared from the body by ordinarybiological processes. In some embodiments, a biodegradable polymercompletely or substantially completely degrades in vivo over the courseof about 30 days or less, where the extent of degradation is based onpercent mass loss of the biodegradable polymer, and wherein completedegradation corresponds to 100% mass loss. Specifically, the mass lossis calculated by comparing the initial weight (W₀) of the polymer withthe weight measured at a pre-determined time point (W_(t)) (such as 30days), as shown in equation (1):

$\begin{matrix}{{{Mass}\mspace{14mu}{loss}\mspace{14mu}(\%)} = {\frac{( {W_{0} - W_{t}} )}{W_{0}} \times 100.}} & (1)\end{matrix}$

In some embodiments, the biodegradable polymer fibers of a meshdescribed herein comprise one or more antimicrobial polymer fibers, suchas one or more chitosan fibers. In other cases, the biodegradablepolymer fibers can include an antimicrobial material dispersed within orbonded to the surface of the fibers. Additionally, in some embodiments,biodegradable polymer fibers described herein include antimicrobialpeptides encapsulated within or bonded to the surface of the fibers.

In some cases, the biodegradable polymer fibers of a wound dressingdescribed herein comprise or are formed from one or more of chitosan,carboxymethyl chitosan (CMC), and polyethylene oxide (PEO) orpolyethylene glycol (PEG). In other instances, the biodegradable polymerfibers of a wound dressing comprise or are formed from an alginate,agarose, starch, polysaccharide, cellulose or cellulose derivative,dextrin, dextran, fibrin, fibrinogen, fibronectin, collagen, gelatin,elastin, laminin, glycosaminoglycan, hyalauronic acid, albumin,polypeptide, polylactic acid (PLA), polyglycolic acid (PGA),polylactic-co-glycolic acid (PLGA), polycaprolactone (PCL),polyglycolide, polyanhydride, polyphosphazene, or polyurethane. Amixture, combination, or copolymer of one or more of the foregoing mayalso be used. For example, in some embodiments, a biodegradable polymerfiber described herein can include a blend of chitosan and PEO. In someinstances, a fiber described herein can be formed from a blend ofchitosan and PEO having a chitosan to PEO ratio between about 1:3 andabout 10:1 by weight or between about 1:1 and about 5:1 by weight. Othercombinations or blends of polymers described herein may also be used toform the biodegradable polymer fibers of a mesh described herein.

Moreover, in some embodiments, the biodegradable polymer fibers of amesh described herein comprise or are formed from one or more polymerscomprising a citrate moiety. A “citrate moiety,” for reference purposesherein, comprises a moiety having the structure of Formula (I):

wherein R₁, R₂, and R₃ are independently —H, —CH₃, —CH₂CH₃, M⁺, or apoint of attachment to the remainder of the polymer;R₄ is —H or a point of attachment to the remainder of the polymer; andM⁺ is a cation such as Na⁺ or K⁺, provided that at least one of R₁, R₂,R₃, and R₄ is a point of attachment to the remainder of the polymer.

For example, in some cases, a polymer of a composition or wound dressingdescribed herein comprises the reaction product of (i) citric acid, acitrate, or an ester of citric acid such as triethyl citrate with (ii) apolyol such as a diol. Non-limiting examples of polyols suitable for usein some embodiments described herein include C2-C20 α,ω-n-alkane diolsor C2-C20 α,ω-alkene diols. In other instances, a polymer of a wounddressing described herein comprises the reaction product of (i) citricacid, a citrate, or an ester of citric acid with (ii) a polyol, and(iii) an amine, an amide, or an isocyanate. An amine, in some cases,comprises one or more primary amines having two to ten carbon atoms. Inother cases, an amine comprises one or more secondary or tertiary amineshaving two to fifteen carbon atoms. An isocyanate, in some embodiments,comprises a monoisocyanate. In other instances, an isocyanate comprisesa diisocyanate such as an alkane diisocyanate. In addition, a polymer ofa wound dressing described herein can also comprise the reaction productof (i) citric acid, a citrate, or an ester of citric acid with (ii) apolyol, and (iii) a polycarboxylic acid such as a dicarboxylic acid or afunctional equivalent of a polycarboxylic acid, such as a cyclicanhydride or an acid chloride of a polycarboxylic acid. Moreover, thepolycarboxylic acid or functional equivalent thereof can be saturated orunsaturated. For example, in some instances, the polycarboxylic acid orfunctional equivalent thereof comprises maleic acid, maleic anhydride,fumaric acid, or fumaryl chloride. In still other embodiments, a polymerdescribed herein comprises the reaction product of (i) citric acid, acitrate, or an ester of citric acid with (ii) a polyol, and (iii) anamino acid such as an alpha-amino acid. An alpha-amino acid, in someembodiments, comprises an L-amino acid, a D-amino acid, or a D,L-aminoacid. In some cases, an alpha-amino acid comprises alanine, arginine,asparagine, aspartic acid, cysteine, glycine, glutamine, glutamic acid,histidine, isoleucine, leucine, lysine, methionine, proline,phenylalanine, serine, threonine, tyrosine, tryptophan, valine, or acombination thereof. Further, in some instances, an alpha-amino acidcomprises an alkyl-substituted alpha-amino acid, such as amethyl-substituted amino acid derived from any of the 22 “standard” orproteinogenic amino acids, such as methyl serine. Additionally, in somecases, an amino acid forms a pendant group or side group of the polymeror oligomer of a composition described herein. Moreover, a reactionproduct of monomers described herein, in some cases, is a condensationreaction product of the monomers. In some cases, a polymer describedherein is a polymer or oligomer described in U.S. Pat. Nos. 8,530,611;8,574,311; or U.S. Pat. No. 8,613,944.

In addition, in some embodiments, a polymer of a wound dressingdescribed herein is formed from one or more monomers of Formula (A) andone or more monomers of Formula (B) or (B′):

wherein R₁, R₂, and R₃ are independently —H, —CH₃, —CH₂CH₃, or M⁺;R₄ is —H;R₅ is —H, —OH, —OCH₃, —OCH₂CH₃, —CH₃, or —CH₂CH₃;R₆ is —H, —CH₃, or —CH₂CH₃;M⁺ is a cation such as Na⁺ or K⁺; andn and m are independently integers ranging from 1 to 20.In some cases, for instance, R₁, R₂, and R₃ are —H, R₅ is —OH, and R₆ is—H.

In some embodiments, a polymer of a wound dressing described herein isformed from one or more monomers of Formula (A), one or more monomers ofFormula (B) or (B′), and one or more monomers of Formula (C):

wherein R₁, R₂, and R₃ are independently —H, —CH₃, —CH₂CH₃, or M⁺;R₄ is —H;R₅ is —H, —OH, —OCH₃, —OCH₂CH₃, —CH₃, or —CH₂CH₃;R₆ is —H, —CH₃, or —CH₂CH₃;M⁺ is a cation such as Na⁺ or K⁺;n and m are independently integers ranging from 1 to 20; andp is an integer ranging from 1 to 10.For example, in some instances, R₁, R₂, and R₃ are —H, or —CH₂CH₃, R₅ is—OH, R₆ is —H, n is 2 to 6, m is 2 to 8, and p is 2 to 6.

Further, in some embodiments of wound dressings described herein, amonomer of Formula (B) or (B′) can be replaced by an alcohol that doesnot have the formula of Formula (B) or (B′). For example, in someembodiments, an unsaturated alcohol or an unsaturated polyol can beused. Moreover, in some cases, a monomer or oligomer of Formula (C) canbe at least partially replaced by an amino acid described herein.

Additionally, a biodegradable polymer described herein can have at leastone ester bond in the backbone of the polymer. In some cases, a polymerhas a plurality of ester bonds in the backbone of the polymer, such asat least three ester bonds, at least four ester bonds, or at least fiveester bonds. In some embodiments, a polymer described herein has betweentwo ester bonds and fifty ester bonds in the backbone of the polymer.

Further, in some cases, a mesh of a wound dressing described herein canbe formed from a mixture or blend of biodegradable polymer fibers formedfrom different polymers described herein.

Moreover, the biodegradable polymer fibers of a mesh described hereincan have any size and shape not inconsistent with the objectives of thepresent disclosure. In some embodiments, for instance, the biodegradablepolymer fibers have an average diameter of about 1000 nm or less. Insome cases, polymer fibers described herein have an average diameterbetween about 10 nm and about 1000 nm or between about 50 nm and about1000 nm. In some cases, polymer fibers described herein can have anaverage diameter between about 10 nm and about 500 nm, between about 10nm and about 100 nm, between about 50 nm and about 500 nm, between about100 nm and about 1000 nm, or between about 500 nm and about 1000 nm. Inother instances, polymer fibers described herein have an averagediameter greater than about 1000 nm. In some embodiments, polymer fibersdescribed herein have an average diameter between about 1000 nm andabout 100 μm, between about 1000 nm and about 10 μm, between about 5 μmand about 100 μm, between about 5 μm and about 50 μm, or between about10 μm and about 100 μm.

Wound dressings described herein, in some embodiments, also comprise afirst active agent dispersed in the biodegradable polymer fibers of amesh. An “active agent,” for reference purposes herein, can comprise anyspecies operable to provide a biological effect when disposed in vivo.Any active agent not inconsistent with the objectives of the presentdisclosure may be used. In some embodiments, the first active agent of awound dressing described herein comprises a growth factor. Any growthfactor not inconsistent with the objectives of the present disclosuremay be used. In some cases, a growth factor described herein canmodulate one or more wound healing processes, such as hemostasis, cellmigration, cell differentiation, ECM formation, and angiogenesis. Insome embodiments, the first active agent comprises an epidermal growthfactor (EGF); a heparin binding EGF; a platelet-derived growth factor(PDGF) such as PDGF-BB; a transforming growth factor beta (TGF-β) suchas TGF-β-1 or TGF-β-2; a vascular endothelial growth factor (VEGF); aninsulin like growth factor (IGF) such as IGF-I or IGF-II; an acidic orbasic fibroblast growth factor (FGF) such as FGF-1 or FGF-2; and/or oneor more isoforms of the foregoing. Other growth factors may also beused.

Moreover, in some embodiments, the identity of the first active agent ofa wound dressing described herein is selected to provide a desired firstbiological effect, such as promotion of blood vessel growth ordevelopment. For example, in some embodiments, the first active agentcomprises a growth factor for angiogenesis and/or the formation ofgranulation tissue, such as VEGF. In other instances, the first growthfactor can comprise a PDGF, and thus may provide tissue inflammationcontrol, granulation, re-epithelialization, and/or remodeling throughouta wound healing process.

A first active agent can be present in a mesh of a wound dressingdescribed herein in any amount not inconsistent with the objectives ofthe present disclosure. For example, in some cases, a first active agentis present in the mesh in an amount up to about 20 weight percent, up toabout 10 weight percent, up to about 5 weight percent, or up to about 1weight percent, based on the total weight of the mesh.

The mesh of a wound dressing described herein can have a variety ofphysical and chemical characteristics. In some embodiments, for example,a mesh of a wound dressing has a high porosity. In some cases, the meshhas a porosity of up to about 90%, up to about 75%, or up to about 50%.In some embodiments, the mesh has a porosity between about 10% and about90%, between about 10% and about 80%, between about 30% and about 90%,or between about 30% and about 70%.

A mesh of a wound dressing described herein can also have a hydrophilicsurface or a hydrophobic surface. The hydrophilicity and/orhydrophobicity of a mesh described herein can be selected based on thechemical composition of one or more of the biodegradable polymer fibersused to form the mesh.

In addition, a mesh of a wound dressing described herein can have anythickness not inconsistent with the objectives of the presentdisclosure. In some cases, the mesh has an average thickness in thez-direction between about 10 nm and about 10 mm, between about 100 nmand about 1 mm, between about 100 nm and about 500 μM, between about 1μm and about 10 mm, between about 1 μm and about 1 mm, between about 10μm and about 10 mm, between about 10 μm and about 1 mm, between about100 μm and about 10 mm, or between about 10 μm and about 1 mm. Otherthicknesses are also possible.

Wound dressings described herein, in some embodiments, also comprise aplurality of biodegradable polymer particles disposed in the mesh of thewound dressing. Such particles can be formed from any biodegradablepolymer not inconsistent with the objectives of the present disclosure.In some cases, the plurality of biodegradable polymer particles areformed from one or more polymers described hereinabove for thebiodegradable polymer fibers of the wound dressing. For example, in someembodiments, the biodegradable polymer particles comprise or are formedfrom one or more of a polyester, polylactic acid, polyglycolic acid,polylactic-co-glycolic acid, polycaprolactone, and copolymers thereof.

The biodegradable polymer particles of a wound dressing described hereincan also have any size and shape not inconsistent with the objectives ofthe present disclosure. In some embodiments, for example, the polymerparticles are nanoparticles having an average size in one, two, or threedimensions of about 1000 nm or less. In some cases, the polymerparticles have an average size in one, two, or three dimensions betweenabout 1 nm and about 1000 nm, between about 1 nm and about 500 nm,between about 10 nm and about 1000 nm, between about 10 nm and about 500nm, between about 10 nm and about 200 nm, between about 50 nm and about1000 nm, between about 50 nm and about 500 nm, between about 100 nm andabout 1000 nm, or between about 100 nm and about 500 nm. In other cases,the biodegradable polymer particles of a wound dressing described hereinare microparticles having a size in one, two, or three dimensionsgreater than 1000 nm. In some embodiments, for instance, the polymerparticles have an average size in one, two, or three dimensions betweenabout 1 μm and about 100 μm, between about 5 μm and about 100 μm,between about 5 μm and about 50 μm, between about 10 μm and about 100μm, between about 10 μm and about 50 μm, or between about 50 μm andabout 100 μm. Moreover, in some cases, the polymer particles of a wounddressing described herein have an average size in one, two, or threedimensions that is smaller than the average diameter of thebiodegradable polymer fibers of the wound dressing.

Further, the polymer particles of a wound dressing described herein canhave a spherical or substantially spherical shape or a polygonal shape.Polymer particles described herein, in some cases, can also berod-shaped. Other shapes are also possible.

Wound dressings described herein, in some embodiments, further comprisea second active agent dispersed in the polymer particles of the wounddressing. The second active agent can comprise any active agent notinconsistent with the objectives of the present disclosure. In somecases, the second active agent is selected to achieve a secondbiological effect, including a second biological effect that maydesirably be temporally coupled with or at least partially separatedfrom a first biological effect. For example, in some cases, the secondbiological effect could be the promotion of wound healing following thepromotion of blood vessel growth. In other cases, the second biologicaleffect could be the promotion of bone growth and development followingthe promotion of blood vessel growth. In some embodiments, the secondactive agent of a wound dressing described herein comprises a growthfactor, including any growth factor described hereinabove for the firstactive agent of a wound dressing. In some cases, for instance, thesecond active agent comprises a growth factor for wound healing or bonegrowth. In other embodiments, the second active agent comprises anosteoinductive growth factor, such as transforming growth factor-β, or abone morphogenetic protein (BMP).

A second active agent can be present in polymer particles of a wounddressing described herein in any amount not inconsistent with theobjectives of the present disclosure. For example, in some cases, asecond active agent is present in the particles in an amount up to about20 weight percent, up to about 10 weight percent, up to about 5 weightpercent, or up to about 1 weight percent, based on the total weight ofthe particles.

In another aspect, wound dressings described herein comprise a stack ofbiodegradable polymer fiber meshes. In some embodiments, the stack ofmeshes is arranged to provide a property gradient in the z-direction,where the z-direction is defined as the stacking direction or height ofthe stack. For example, in some cases, the meshes are porous meshes andthe stack of meshes exhibits a porosity gradient in the z-direction. Insome such embodiments, the porosity of the stack decreases from thebottom to the top of the stack. Alternatively, in other instances, theporosity of the stack increases from the bottom to the top of the stack.

Further, in some embodiments, one or more of the meshes of a stackdescribed herein are perforated meshes and the stack of meshes exhibitsa perforation gradient in the z-direction. In some such cases, thedegree of perforation of the meshes decreases from the bottom to the topof the stack. In other instances, the degree of perforation of themeshes increases from the bottom to the top of the stack. Moreover, insome embodiments, the top mesh of a stack of meshes described herein isnon-perforated, where the “top” mesh refers to the mesh farthest fromthe side of the stack to be placed in contact with a wound.

The “degree of perforation” of a mesh described herein can be based onthe number of perforations or holes per unit area of a surface of themesh, the average size of the perforations or holes, or the total areaor volume of the perforations or holes. The “degree of perforation” canalso be based on the shapes of the perforations or holes. In someembodiments, the perforations or holes of a perforated mesh describedherein have an average size or diameter of at least about 10 μm, atleast about 20 μm, at least about 30 μm, at least about 50 μm, at leastabout 100 μm, or at least about 150 μm. In some cases, the perforationsor holes of a perforated mesh described herein have an average size ordiameter between about 10 μm and about 10 mm, between about 10 μm andabout 5 mm, between about 10 μm and about 1 mm, between about 50 μm andabout 10 mm, between about 50 μm and about 5 mm, between about 50 μm andabout 1 mm, between about 100 μm and about 10 mm, between about 100 μmand about 5 mm, or between about 100 μm and about 1 mm. Perforationshaving such sizes, in some cases, can provide sufficient space forregenerated tissue to grow into and penetrate the mesh during woundhealing. Further, in some instances, the perforations or holes of aperforated mesh described herein have a pitch or average distancebetween perforations or holes of about 0.1 mm to about 10 mm or about0.5 mm to about 5 mm, where the average distance between perforations orholes is based on the center-to-center distance between adjacentperforations or holes. In addition, in some embodiments, a perforatedmesh described herein has a perforation or hole density of at leastabout 10 perforations/cm², at least about 20 perforations/cm², at leastabout 30 perforations/cm², at least about 50 perforations/cm², or atleast about 70 perforations/cm², where the area is based on the totalarea of a perforated surface of the perforated mesh and where theaverage size of the perforations can be a size described herein, such asa size greater than about 50 μm. In some cases, a perforated meshdescribed herein has a perforation or hole density between about 10perforations/cm² and about 200 perforations/cm², between about 20perforations/cm² and about 150 perforations/cm², or between about 30perforations/cm² and about 100 perforations/cm², wherein the averagesize of the perforations is a size described hereinabove.

A stack of perforated meshes having a porosity and/or perforationgradient described herein, in some cases, can permit improvedpenetration of cells from a wound site into the stack. In addition, astack of meshes having a porosity and/or perforation gradient describedherein, in some embodiments, can permit the gradual transfer ofmechanical load from the stack of meshes to the tissue of the wound siteas healing occurs. Specifically, wound dressings comprising a stack ofmeshes described herein, in some embodiments, can provide gradualtransfer of mechanical loads from the mesh itself to regeneratedbiological tissue as wound healing progresses. Not intending to be boundby theory, it is believed that such gradual transfer of mechanical load,in some cases, can be achieved by the cell penetration afforded byperforations and perforation gradients described herein. Additionally,in some embodiments, a perforated mesh described herein can exhibit oneor more mechanical properties provided in Table I below, when measuredaccording to ASTM D412A.

TABLE I Mechanical Properties of Perforated Meshes. Elongation atInitial Modulus Peak Stress Break (%) (MPa)(MPa) >300 >2 >1 >400 >5 >2 >500 >10 >3  50-600 2-15 1-6  50-500 2-101-5 100-500 5-15 1-4 200-600 5-10 2-4 300-600 10-15  3-5

A wound dressing comprising a stack of meshes described herein caninclude any number of meshes not inconsistent with the objectives of thepresent disclosure. In some cases, for example, a wound dressingcomprises at least two, at least three, at least five, at least 10, orat least 20 meshes in a stacked configuration. In some instances, awound dressing comprises up to 50 or up to 100 meshes in a stackedconfiguration. Further, the meshes of such a wound dressing canindependently have any structure or property of a mesh described herein.

For example, in some embodiments, a wound dressing described hereincomprises a first perforated mesh formed from a first plurality ofbiodegradable polymer fibers; and a second perforated mesh formed from asecond plurality of biodegradable polymer fibers, wherein the secondperforated mesh is disposed on the first perforated mesh in a stackedconfiguration and the first perforated mesh has a higher degree ofperforation than the second perforated mesh. Moreover, in some cases, awound dressing further comprises a third perforated mesh formed from athird plurality of biodegradable polymer fibers, wherein the thirdperforated mesh is disposed on the second perforated mesh in a stackedconfiguration and the third perforated mesh has a higher degree ofperforation than the first perforated mesh and the second perforatedmesh. Additionally, if desired, wound dressings described herein canfurther comprise additional perforated meshes or non-perforated meshesin a stacked configuration.

For example, in some embodiments, the top mesh of a wound dressingdescribed herein is a non-perforated mesh. Thus, in some cases, a wounddressing described herein further comprises a fourth mesh formed from afourth plurality of biodegradable polymer fibers, wherein the fourthmesh is disposed on the third perforated mesh in a stacked configurationand the fourth mesh is non-perforated or has a lower degree ofperforation than the third perforated mesh. A wound dressing having sucha structure, in some cases, can provide a physical barrier to completetissue penetration of the wound dressing on the side of the wounddressing farther from the wound.

As described above, it is generally to be understood that the number ofstacked meshes in a wound dressing described herein is not particularlylimited. Instead, any desired number of meshes can be used to provide awound dressing having a desired thickness and/or a desired propertygradient in the z-direction. The meshes of a stack described herein canalso be arranged to provide a wound dressing having regularly orirregularly repeating properties in the z-direction. In some cases, forinstance, the meshes of a stack described herein have alternatinghydrophobicity and hydrophilicity. Thus, in some embodiments, a wounddressing described herein comprises a first perforated mesh and a secondperforated mesh in a stacked configuration, wherein the first perforatedmesh is hydrophilic and the second perforated mesh is hydrophobic. Otherarrangements of meshes are also possible.

Additionally, one or more meshes of a stack described herein can have astructure described hereinabove for wound dressings comprising activeagents. For example, in some cases, one or more active agents aredispersed in the biodegradable polymer fibers of a first perforated meshand/or a second perforated mesh of a stack described herein. Further, insome embodiments, a plurality of biodegradable polymer particles isdisposed in a first perforated mesh and/or a second perforated mesh of awound dressing described herein. Moreover, in such cases, one or moresecond active agents can be dispersed in the biodegradable polymerparticles. The first and second active agents of such a wound dressingcan comprise any first active agent, second active agent, andcombination of first and second active agents described hereinabove. Insome embodiments, for instance, the in vivo or in vitro release profileof a first active agent of a stack differs from the in vivo or in vitrorelease profile of a second active agent of the stack. In such cases,the in vivo or in vitro release profiles of the active agents can differin a manner described hereinabove. For example, in some cases, the invivo or in vitro release profile of the first active agent and the invivo or in vitro release profile of the second active agent of a stackof meshes described herein overlap by less than about 70%. In othercases, the release profiles are entirely non-overlapping.

In addition, wound dressings described herein, in some embodiments, canalso include one or more electrically conductive components for neuraland muscular tissue engineering applications. For example, in somecases, the mesh of a wound dressing described herein further comprisesone or more electrically conductive polymer fibers, such as one or morefibers formed from polypyrrole, polyaniline, or a polythiophene such aspoly(3,4-ethylendioxythiophene) (PEDOT). Similarly, in some instances, awound dressing described herein can comprise a plurality of electricallyconductive polymer particles, such as polyaniline particles, polypyrroleparticles, or PEDOT particles. Other electrically conductive polymerfibers and particles can also be used. Such electrically conductivepolymer fibers and particles can be used in addition to the componentsof biodegradable wound dressings described herein or in place of suchcomponents. For example, in some instances, a wound dressing describedherein comprises a mesh formed from a plurality of electricallyconductive polymer fibers and a plurality of electrically conductivepolymer particles disposed in the mesh. The particles can be disposedbetween the fibers of the mesh or within the fibers of the mesh.Further, in some cases, one or both of the electrically conductivepolymer fibers and the electrically conductive polymer particlescomprise an active agent. In some embodiments, for instance, theelectrically conductive polymer fibers and the electrically conductivepolymer particles comprise a combination of first and second activeagents described herein.

Various components of compositions and wound dressings have beendescribed herein. It is to be understood that a composition or wounddressing according to the present disclosure can comprise anycombination of components and features not inconsistent with theobjectives of the present disclosure. For example, in some cases, awound dressing described herein comprises any mesh described herein incombination with any polymer particles described herein and any activeagents described herein.

Wound dressings having a structure described hereinabove can be made inany manner not inconsistent with the objectives of the presentdisclosure. For example, in some embodiments, a wound dressing describedherein is made by an electrospinning process. In some cases, such amethod of making a wound dressing comprises electrospinning a mixturecomprising a first biodegradable polymer, a first solvent, and a firstactive agent such as a first growth factor. Electrospinning such amixture can provide a plurality of polymer fibers formed from the firstbiodegradable polymer, wherein the first active agent is dispersedwithin the polymer fibers. Further, as described further hereinbelow,the electrospinning process can form the polymer fibers into a non-wovenmesh. In addition, in some embodiments, the mixture for electrospinningcan further comprise a plurality of biodegradable polymer particlesdescribed herein. Electrospinning such a mixture can provide a meshdescribed herein, wherein the polymer particles are dispersed within thepolymer fibers. In some such cases, the solvent of the mixture isselected for its compatibility with the biodegradable polymers of thefibers and/or particles and for its compatibility with the first and/orsecond active agents. For example, in some embodiments, the firstbiodegradable polymer is soluble in the solvent of the mixture but thebiodegradable polymer particles are not soluble in the solvent. In somesuch cases, for instance, an aqueous solvent is used with awater-soluble first biodegradable polymer and with hydrophobic polymerparticles.

Alternatively, a plurality of biodegradable polymer particles can bedisposed in between fibers of a mesh by forming a mesh of polymer fibersin a manner described hereinabove, followed by treating the mesh with asolution or mixture comprising the polymer particles. For example, insome cases, a particle solution or mixture can be drop cast onto themesh. The mesh can also be immersed in the particle solution or mixture.

Similarly, electrospinning may also be used to provide a wound dressingcomprising a stack of meshes described herein. Such a method, in someembodiments, comprises forming a plurality of meshes in a mannerdescribed hereinabove and then stacking the meshes. In addition, in somecases, one or more of the meshes are perforated before or after stackingthe meshes to provide the wound dressing. As described furtherhereinbelow, multiple meshes can be stacked in any manner notinconsistent with the objectives of the present disclosure. In somecases, for example, multiple meshes can be stacked by directlyelectrospinning different meshes one on top of the other, by physicallypressing meshes together, by applying a biodegradable adhesive betweenadjacent meshes, or by applying mild solvent for surface weldingadjacent meshes. Other stacking techniques may also be used.

II. Methods of Treating a Wound

In another aspect, methods of treating a wound are described herein. Insome embodiments, a method of treating a wound comprises applying acomposition or wound dressing described herein to a surface of a wound.Any composition or wound dressing described hereinabove in Section I maybe used. In some cases, for instance, a wound dressing comprises a meshformed from a plurality of biodegradable polymer fibers; a first activeagent dispersed in the biodegradable polymer fibers; a plurality ofbiodegradable polymer particles disposed in the mesh; and a secondactive agent dispersed in the biodegradable polymer particles. Further,in some instances, the particles are disposed within the fibers of themesh. Additionally, a method comprising the application of such a wounddressing, in some cases, can further comprise at least partiallydegrading the biodegradable polymer fibers to release the first activeagent into the wound. Degrading the polymer fibers, in some cases,comprises cleaving one or more chemical bonds such as one or more esterbonds in the polymer fibers. Moreover, degrading the polymer fibers of awound dressing can, in some embodiments, provide an in vivo releaseprofile of the first active agent that corresponds to an in vivo releaseprofile described hereinabove in Section I.

In addition, in some cases, a method described herein further comprisesat least partially degrading the biodegradable polymer particles torelease the second active agent into the wound. Degrading the polymerparticles can comprise cleaving one or more chemical bonds in theparticles, including one or more ester bonds. Further, degrading thepolymer particles of a wound dressing in a manner described herein canprovide an in vivo release profile of the second active agent thatcorresponds to an in vivo release profile described hereinabove inSection I. In some cases, for instance, the second active agent isreleased from the wound dressing after the first active agent isreleased from the wound dressing. Thus, as described further herein, amethod of treating a wound described herein can comprise using a singlewound dressing to provide a plurality of active agents to a wound sitein a temporally controlled and/or bifurcated manner. For example, insome cases, the first active agent of a method described hereincomprises a growth factor for angiogenesis, and the second active agentof the method comprises a growth factor for wound healing or bonegrowth.

In other embodiments of methods described herein, the composition orwound dressing applied to a wound comprises a stack of meshes. Such acomposition or wound dressing can comprise any wound dressing describedhereinabove in Section I. For example, in some instances, the wounddressing comprises a first perforated mesh formed from a first pluralityof biodegradable polymer fibers; and a second perforated mesh formedfrom a second plurality of biodegradable polymer fibers, wherein thesecond perforated mesh is disposed on the first perforated mesh in astacked configuration and the first perforated mesh has a higher degreeof perforation than the second perforated mesh.

In addition, a method described herein can be used to treat any type ofwound not inconsistent with the objectives of the present disclosure. Insome embodiments, for instance, the wound comprises a skin wound. Insome cases, the wound comprises a diabetic ulcer or hernia.

Some embodiments described herein are further illustrated in thefollowing non-limiting examples.

Example 1 Wound Dressings

Wound dressings according to some embodiments described herein wereprovided and used to treat wounds as follows.

Materials

Chitosan (CS, medium molecular weight, 75-85% deacetylated),polyethylene oxide (PEO, M_(n)=600,000 Dalton), bovine serum albumin(BSA), acetic acid, and chloroform were purchased from Sigma Aldrich(St. Louis, Mo.). Poly-lactic-co-glycolic acid (PLGA) (50:50) waspurchased from Lakeshore Biomaterials (Birmingham, Ala.). PlateletDerived Growth Factor-BB (PDGF-BB, Human Recombinant) and VascularEndothelial Growth Factor (VEGF, Rat Recombinant) were purchased fromProspec (East Brunswick, N.J.). HDF (Adult Human Dermal Fibroblast)cells were purchased from ATCC (Manassas, Va.). Gram-negativeEscherichia coli (E. coli, 25922™) and gram-positive Staphylococcusaureus (S. aureus, 25923™) were also obtained from ATCC.

Biodegradable Polymer Particles

PLGA nanoparticles were fabricated using the double-emulsion techniquedescribed by Menon et al., “Effects of surfactants on the properties ofPLGA nanoparticles,” Journal of Biomedical Materials Research, Part A,2012, 100, 1998-2005. Briefly, 200 μL of 5% w/v BSA or 2% w/v PDGF-BBaqueous solution was added to 3.33 mL of 3% w/v PLGA aqueous solutionand sonicated at 30 W for 2 minutes. This o/w solution was then addeddropwise to 12 mL 2% PVA solution and sonicated at 20 W for two minutes.This final w/o/w solution was then de-solvated overnight using amagnetic stirrer. Centrifugation was then performed at 4,000 rpm for 5minutes to remove particle aggregates. The BSA or PDGF-BB loaded PLGAnanoparticles were obtained via freeze-drying. In addition, thesupernatant from the nanoparticle formation process was also collectedto determine the loading efficiency.

Biodegradable Polymer Mesh

A polymer mesh was prepared by electrospinning. First, stock polymersolutions were prepared. Specifically, a solution of chitosan (CS) at aconcentration of 2.5% w/v was prepared in 90% acetic acid. A solution ofPEO at a concentration of 8% w/v was prepared in deionized (DI) water atroom temperature. Next, two CS/PEO blend solutions were prepared bymixing the two stock solutions at 1:1 and 2:1 chitosan to PEO volumeratios. For reference purposes, polymer fibers formed from thesemixtures, without nanoparticles, are denoted as 1:1 CS/PEO and 2:1CS/PEO, respectively. To provide polymer fibers comprisingnanoparticles, 20% by weight of PLGA nanoparticles (based on the weightof PEO) was added to the CS/PEO mixtures and sonicated for 10-15 minutesat 20 W to obtain complete or substantially complete dispersion of thenanoparticles. These fibers were denoted as 1:1 CS/PEO-NPs and 2:1CS/PEO-NPs.

For electrospinning, each of the blended solutions above wasindividually loaded into a 5 mL syringe equipped with an 18-gauge bluntneedle tip. For each electrospinning experiment, the syringe was loadedinto a syringe pump. The contents of the syringe were delivered forelectrospinning by driving the syringe plunger with the syringe pump ata flow rate of 1.5 μL/min. The tip of the syringe was disposed 15 cmaway from an aluminum mesh collector, and a DC voltage of 18 kV wasapplied between the collector and the tip. All electrospinningexperiments were carried out at ambient temperature (about 25° C.) and arelative humidity of 15-20%.

Results

The surface morphology of the electrospun nanofiber mesh wascharacterized using a scanning electron microscope (SEM) (Hitachi,S-3000N). All samples were first sputter-coated with silver. Fiberdiameters were determined from SEM images using Image-J software. Foreach mesh, 100 fibers were considered from three different images tocalculate the average diameter. To visualize the nanoparticles withinthe nanofibers, indocyanine green (ICG) loaded PLGA nanoparticles wereprepared and electrospun. Fluorescent images were captured using afluorescence microscope equipped with a TRITC filter.

To assess the active agent release kinetics, BSA was selected as a modelprotein. Specifically, BSA was incorporated into the biodegradablepolymer fibers and/or the biodegradable polymer particles in the mannerdescribed above. Meshes containing BSA and weighing 10.0-11.0 mg wereloaded into 100-kDa dialysis membranes and placed in 0.1 M phosphatebuffered saline (PBS) solutions. The samples were then placed on anorbital shaker at 37° C. At predetermined time points, 1 mL of PBSsolution was collected and replaced with 1 mL fresh PBS. The releaseprofile of BSA (60 kDa), either from the nanofibers themselves or fromPLGA nanoparticles within the nanofibers, was analyzed using standardBSA protein assays following the manufacturer's instructions. Cumulativerelease over a period of 21 days was performed on all samples.

Adult Human Dermal Fibroblasts (HDFs) were cultured in completeDulbecco's Modified Eagle's medium (DMEM) with supplements of 10% FetalBovine Serum (FBS) and 1% penicillin/streptomycin solution. Cells weresub-cultured until approximately 80% confluency and maintained at ahumidified atmosphere of 95% air and 5% CO₂. For in vitro cellproliferation on nanofiber meshes, mesh samples (3 mm in diameter) werevacuum dried overnight and then UV-sterilized for 1 hour. Samples werethen placed in a 96-well plate and seeded with 5000 cells/well. A tissueculture plate was used as a control. MTS assays were performed at timepoints of 1, 3, 5, and 7 days following seeding. Absorbance at 490 nmwas measured, and the cell proliferation was plotted over time as apercentage over the control sample at day 1.

To assess the antibacterial activity of the samples, three types ofnanofiber meshes were used. Specifically, 1:1 CS/PEO, 2:1 CS/PEO, and2:1 CS/PEO-NPs were used. All mesh samples were vacuum-dried and UVsterilized. 20 mg of each type of mesh were used. E. coli and S. aureuswere reconstituted based on the supplier's instructions. A bacteriasuspension and PEO nanofibers without chitosan were chosen as thenegative controls for antibacterial activity. Penstrep was used as thepositive control. For each sample, a bacterial suspension was preparedat an optical density (OD) of 0.011 at 600 nm (measured by a UV-visspectrophotometer), added to the sample, and incubated at 37° C. Afterincubation, the absorbance at 600 nm of each sample was measured atpredetermined time points. The average of background samples wassubtracted from the test samples and plotted over time. All operationswere carried out in aseptic conditions.

Sprague-Dawley rats weighting approximately 250 g were used for in vivostudies. Specifically, full thickness skin wound healing studies werecarried out. All animals were treated and used in accordance with theprotocol approved by the University of Texas at Arlington Animal Careand Use Committee (IACUC). Animals were anesthetized with ketamine (40mg/kg) and xylazine (5 mg/kg), and then shaved on the back. A 5 mmdiameter biopsy puncher was used to create a wound along the dorsal sideof the skin. Four wounds were created on each rat, and then controls(open wound and Hydrofera Blue®) and the test samples were placed on thewound site randomly. Changes in the wound areas were measured using acaliper at 1, 7, 14, and 28 days after initial wounding and placement ofthe wound dressings. At each time point, the surrounding skin and muscleincluding wound areas were removed and fixed by 10% neutral buffedformalin. Tissue samples were embedded in paraffin and sectioned.Hematoxylin-eosin (H&E) and Masson's Trichrome staining were performedto evaluate the skin tissue sections.

Physical measurements of surface epidermal tongue and granulation tissuethickness of the H&E images were measured using Image-J. Collagenquantification was carried out by measuring the blue area percentage ofthe wound area with Masson's Trichrome staining.

All data herein is presented as the mean±standard deviation (SD).Statistical analysis of all data was performed using 1-way ANOVA(StatView), where p values <0.05 were considered statisticallysignificant (n=6).

FIG. 1 is a schematic illustration of an exemplary wound dressing (100)described herein and a method of treating a wound (200) using the wounddressing (100). As illustrated in FIG. 1, PDGF-BB (140) was encapsulatedwithin PLGA nanoparticles (130) (average diameter of 153±36 nm, asdetermined by Dynamic Light Scattering) of the wound dressing (100), andthen dispersed in CS/PEO nanofibers (110). In addition, VEGF (120) wasloaded into the nanofibers (110) of the mesh. The wound dressing (100)was then applied to the wound (200). Following application of the wounddressing (100) to the wound (200), the relatively fast-releasing VEGF(120) and the relatively slow- or sustained-releasing PDGF-BB (140) werereleased into the wound (200) to promote wound healing.

FIG. 2 illustrates SEM images of various meshes described herein havingthe structure illustrated in FIG. 1. FIG. 2A corresponds to 2:1CS/PEO-NPs. FIG. 2B corresponds to 1:1 CS/PEO-NPs. The SEM images showsmooth, uniform, and beadless fibrous nonwoven structures. The 1:1CS/PEO-NPs mesh had a smaller average fiber diameter of 116±39 nm, whilethe 2:1 CS/PEO-NPs mesh had an average fiber diameter of 132±39 nm. Inorder to visualize the nanoparticles within the fibers, PLGAnanoparticles were loaded with indocyanine green (ICG) and imaged byfluorescence microscopy. FIG. 2C is a fluorescence image of ICG loadedNPs in CS/PEO fibers. As shown in FIG. 2C, the nanoparticles werelocated within fibers and were uniformly distributed. FIG. 2Dillustrates diameter distributions of the electrospun fibers of thesamples.

FIG. 3 illustrates BSA release kinetics from nanofibers andnanoparticles within fibers. For reference purposes, the nomenclature“Fiber” and “NPs” is used to indicate which portion of the mesh wasloaded with BSA. For example, the 2:1 CS/PEO-Fiber release profilecorresponds to BSA loaded into the nanofibers only, and the 1:1CS/PEO-NPs and 2:1 CS/PEO-NPs release profiles correspond to BSA loadedinto PLGA nanoparticles that were encapsulated in the nanofibers, andwherein no BSA was dispersed in the nanofibers themselves. Asillustrated in FIG. 3, BSA was released from nanofibers quickly. Forexample, the 2:1 CS/PEO-Fiber release profile included an initial burstrelease of 64% within the first 30 minutes. The BSA loaded within the2:1 CS/PEO mesh was all released by day 3. In contrast, BSA releasedfrom PLGA nanoparticles within a 2:1 CS/PEO-NPs mesh showed only a smallinitial burst release of 16% at day 1. In addition, BSA release fromPLGA nanoparticles for both meshes exhibited a sustained releasepattern.

FIG. 4 illustrates the results of cell proliferation experiments for thefollowing sample meshes: 1:1 CS/PEO, 2:1 CS/PEO, and 2:1 CS/PEO-NPs(PLGA nanoparticles loaded with PDGF-BB). Meshes were seeded with HDFsand MTS assay was used to quantify the cell viability (* p<0.01). All ofthe PEO/CS meshes were cytocompatible throughout the time period of theexperiment and exhibited more cell growth than the control. Cellproliferation was significantly increased on days 5 and 7 on all meshescompared to the control. A growth of 116.9±2.9% was observed on 1:1CS/PEO on day 5, and 115.2±2.8% growth was observed on the 2:1 CS/PEOmesh. On day 7, the 1:1 CS/PEO growth was 132.6±1.8%, and aproliferation of 132.5±2.9% was observed for 2:1 CS/PEO. The CS/PEO-NPssample with PDGF-BB loaded nanoparticles exhibited significantly fastercell growth for day 5 (140.9±0.8%) and day 7 (156.8±6.6%) compared tothe tissue culture plate control.

FIG. 5 illustrates antibacterial properties of various CS/PEO-NPs meshescompared to negative controls (cell suspension and PEO mesh) and apositive control (Penstrep solution). Antibacterial activity wasassessed based on bacterial optical density, as described above. Twotypes of bacteria, E. coli (FIG. 5A) and S. aureus (FIG. 5B) were used.As illustrated in FIG. 5, the negative controls showed continuousexpansion of both E. coli and S. aureus. In contrast, 1:1 CS/PEO-NPs and2:1 CS/PEO-NPs meshes exhibited antibacterial activity against both E.coli and S. aureus compared to negative controls (* p<0.05).

FIG. 6 illustrates wound healing properties of various meshes describedherein. Specifically, FIG. 6A illustrates representative macroscopicappearances of wound closures at 0, 1, 2, and 4 weeks after treatment ofskin wounds. Electrospun 2:1 CS/PEO-NPs without growth factor (denotedas 2:1 CS/PEO in FIGS. 6-8) and 2:1 CS/PEO-NPs with VEGF in the fibersand PDGF-BB in PLGA nanoparticles (denoted as 2:1 CS/PEO-NPs in FIGS.6-8) were placed and adhered on the wound site easily. Further, comparedto commercial Hydrofera Blue, which requires biological adhesives to befixed on a wound site, the meshes were much easier to attach to wounds.In addition, approximately 4 hours after placement, the meshes becameinvisible to the eye. At 1 week after treatment, no infection wasobserved for all samples. Higher granulation and regenerated epidermiswere observed for 2:1 CS/PEO-NPs meshes, as confirmed later byhistological analysis. At 2 weeks after treatment, scabs fell from theskin wounds for all samples. Again, 2:1 CS/PEO-NPs samples exhibitedfaster healing with more regenerated skin and more hair growth. After 4weeks, all wounds appeared to be closed. Scabs were observed onHydrofera Blue samples only.

FIG. 6B illustrates quantitative measurements of wound size reduction orwound closure as a function of time (* p<0.01). As illustrated in FIG.6B, wound areas for 2:1 CS/PEO-NPs meshes were significantly smallerthan those of other samples at weeks 1 and 2 (p<0.05). It was alsoobserved that at week 1 Hydrofera Blue exhibited a slightly increasedwound size due to extensive scar formation. After 4 weeks of treatment,all wounds were closed. 2:1 CS/PEO-NPs exhibited the smallest scarformation area and the greatest hair coverage.

FIG. 7 illustrates the results of histological evaluation of the woundstreated by CS/PEO-NPs meshes and Hydrofera Blue wound dressing. FIG. 7Aillustrates H&E staining for skin wound samples of control (open wound),2:1 CS/PEO, 2:1 CS/PEO-NPs, and Hydrofera Blue samples after 1 and 2weeks of treatment. FIG. 7B illustrates epithelial tongue length after 1week of treatment. FIG. 7C illustrates the capillary density at woundsites after 1 and 2 weeks of treatment. FIG. 7D illustrates granulationtissue thickness after 1 and 2 weeks of treatment. (*, ** p<0.05).Longer epithelial tongues were observed for 2:1 CS/PEO-NPs samples (FIG.7B). At one week and two weeks, significantly more newly formedcapillaries within the wound site were observed for 2:1 CS/PEO-NPscompared to open wound (p<0.01) (FIG. 7C). After 2 weeks of treatment,full coverage of new epithelium was identified for all samples exceptthe Hydrofera Blue samples. In addition, with a complete closure ofepithelium, rapid clearance of PEO, and sustained release of PDGF-BB,the granulation tissue thickness for 2:1 CS/PEO-NPs at week 2 wassignificantly reduced compared to that of week 1 and open wound control(p<0.01), suggesting a transition from Phase I (inflammation) to PhaseII (proliferation) of the healing process (FIG. 7D). The control andHydrofera Blue samples exhibited thicker layers of granulation.

Masson's Trichrome staining was performed to assess the collagendeposition (blue) in the wound site. FIG. 8 illustrates collagenstaining images and quantification of wounds treated by CS/PEO-NP meshesand Hydrofera Blue wound dressing. Specifically, FIG. 8A illustratesMasson's Trichrome staining of the control, 2:1 CS/PEO, 2:1 CS/PEO-NPs,and Hydrofera Blue samples at 2 and 4 weeks post-treatment. FIG. 8Billustrates collagen quantification of each wound area at 2 weeks, andFIG. 8C illustrates collagen quantification of each wound area at 4weeks after treatment (* p<0.05). 2:1 CS/PEO showed a significantly(p<0.05) higher amount of collagen deposition at 2 weeks aftertreatment. A higher amount of myofibroblast formation at the wound sitewas also identified in the 2:1 CS/PEO-NPs samples compared to the openwound. Compared to open wound control and nanofibers without growthfactors, more mature collagen fibers were observed for 2:1 CS/PEO-NPssamples with a lower inflammatory cell presence. More collagen tissuecould help the reconstruction of ECM and further support skin tissuegrowth. After 4 weeks of treatment the growth factor-releasing meshesexhibited the lowest collagen content at the wound area. Not intendingto be bound by theory, it is believed that this observation may be dueto more mature collagen formation and increased hair follicleregeneration. Further, such morphology could indicate that a remodelingphase was already reached for 2:1 CS/PEO-NPs at 4 weeks, while othersamples still remained at the tissue regeneration phase.

Example 2 Wound Dressings Comprising Stacks of Meshes

Wound dressings comprising stacks of meshes according to someembodiments described herein were prepared as follows. First,biodegradable meshes including polymers such as PLA, PLGA, PCL,collagen, hyaluronic acid (HA), gelatin, polyethylene oxide (PEO),chitosan, and carboxylmethyl chitosan (CMC) were obtained byelectrospinning in a manner described hereinabove. Next, a micro-needlearray with various needle sizes and densities was used to punch orperforate individual electrospun meshes to provide micro-holes orperforations through the individual electrospun meshes. Different sizesand densities of holes or perforations could be patterned on bothhydrophobic and hydrophilic meshes. Following fabrication of individualperforated meshes, multiple meshes made of identical or differentmaterials were stacked together to create a mesh assembly with graduallydecreasing perforation sizes and/or densities from one side to theother. Such a structure allowed cells to penetrate from one side of themesh assembly to the other side gradually. The top layer (the sideopposite the wound in this Example) of each stack was formed from anon-perforated mesh. The top layer thus formed a physical barrier fortissue penetration. As described herein, such wound dressings could beused as hernia meshes for hernia repair applications.

FIG. 9 illustrates the foregoing fabricating and assembling steps.Specifically, in the embodiment of FIG. 9, a stack (300) of meshes (310,320, 330, 340, 350) is formed by electrospinning and perforating themeshes (310, 320, 330, 340, 350) individually. For perforation, amicro-patterned perforation or punch apparatus (400) is used to provideperforations or holes (311, 321, 331) in three of the meshes (310, 320,330). The other two meshes (340, 350) are not perforated. Theperforation or punch apparatus (400) comprises an array of needles (410)that can vary in needle density and/or needle size. As illustrated inFIG. 9, only the perforation of the top mesh (310) is shown. Further, inthe embodiment of FIG. 9, the meshes (310, 320, 330, 340, 350) form aperforation gradient in the z-direction, where the degree of perforationdecreases from the top mesh (310) toward the bottom mesh (350). Inaddition, in the embodiment of FIG. 9, the meshes (310, 320, 330, 340,350) are arranged in the stack (300) in an alternating hydrophilic andhydrophobic manner. In particular, hydrophilic meshes (320, 340)alternate with hydrophobic meshes (310, 330, 350) in the stack (300).

Exemplary perforated meshes are illustrated in FIG. 10. Specifically,FIG. 10 illustrates SEM images of perforated electrospunpolycaprolactone (PCL) meshes. FIGS. 10A and 10B are low magnificationimages. FIG. 10C is a high magnification image of a single perforationof a perforated PCL mesh. As shown in FIGS. 10A-C, the micro-needlearray punched clean-cut perforations with a diameter of 150 μm and apitch between holes of 1 mm. Such perforations could provide ampleaccess for tissue ingrowth, unlike some other electrospun meshes whosepores are too small for cell/tissue penetration. FIG. 10D is across-sectional image of a 3-layer stack of meshes comprising a PCL meshdisposed in between two PEO/CMC meshes. The PEO/CMC meshes could beeasily hydrated and intimately attached to the sandwiched PCL mesh.

The mechanical performance of perforated and non-perforated meshes wasevaluated by tensile tests. Specifically, mesh samples cut into stripswith a width of 5 mm and a length of 30 mm were tested. The gaugebetween grips was 10 mm. The crosshead elongation speed was 100 mm/min.Some results for PCL meshes with different degrees of perforation areillustrated in FIG. 11 (*, p<0.01). Perforation densities for the mesheswith 1 mm diameter perforations and 0.15 mm diameter perforations are 36and 64 perforations/cm², respectively. FIG. 11A illustrates the peakstress of the meshes. FIG. 11B illustrates the initial (Young's) modulusof the meshes. FIG. 11C illustrates the elongation at break of themeshes. FIG. 11D illustrates a representative stress-strain curve ofeach sample. As illustrated in FIG. 11, The peak stress and elongationat break decreased significantly after perforation of the meshes.However, such perforated meshes were also shown to be suitable forhernia repair applications. In addition, the mechanical properties ofmeshes described herein, in some cases, can be tuned by varying materialcompositions, electrospinning conditions, and/or degree of perforation.

Various embodiments of the present invention have been described infulfillment of the various objectives of the invention. It should berecognized that these embodiments are merely illustrative of theprinciples of the present invention. Numerous modifications andadaptations thereof will be readily apparent to those skilled in the artwithout departing from the spirit and scope of the invention.

That which is claimed is:
 1. A wound dressing comprising: a plurality ofmeshes formed from a plurality of biodegradable polymer fibers; whereinthe biodegradable polymer fibers comprise a first active agent; whereinthe plurality of meshes further comprise a plurality of biodegradablepolymer particles; wherein the biodegradable polymer particles comprisea second active agent; wherein an in vivo or in vitro release profile ofthe first active agent differs from an in vivo or in vitro releaseprofile of the second active agent; such that the in vivo or in vitrorelease profile of the first active agent and the in vivo or in vitrorelease profile of the second active agent overlap by less than about70%; wherein the biodegradable polymer fibers have an average diameterof between about 10 nm and about 500 nm; wherein the biodegradablepolymer particles have an average size in one, two or three dimensionsof between about 10 nm and about 200 nm; wherein the plurality of meshesare arranged in a stack wherein the stack has a top surface and a bottomsurface, wherein the bottom surface is defined by a mesh placed on awound and the top surface is defined by a mesh disposed farthermost fromthe wound, wherein the stack having a property gradient in a z-directionof the stack such that the property increases from the bottom surface tothe top surface or such that the property decreases from the bottomsurface to the top surface of the stack, wherein the z-direction isdefined in a stacking direction; and wherein the wound dressing providesa fibrous scaffold for supporting cell growth; wherein the plurality ofmeshes is at least three meshes, and wherein the property is defined bya mesh porosity or a mesh perforation.
 2. The wound dressing of claim 1,wherein the plurality of meshes comprises a first perforated mesh and asecond perforated mesh arranged in the stack, the first mesh having agreater degree of perforation than the second mesh.
 3. The wounddressing of claim 2, wherein the first perforated mesh has an averagesize perforation size between 10 μm to about 10 mm.
 4. The wounddressing of claim 2, wherein the first perforated mesh has a perforationdensity between about 10 perforations/cm² to about 200 perforations/cm².5. The wound dressing of claim 1, wherein the plurality of meshescomprises a first porous mesh and a second porous mesh arranged in thestack, the first mesh having a greater degree of porosity than thesecond mesh, and wherein the first porous mesh has a porosity betweenabout 10% to about 90%.
 6. The wound dressing of claim 1, wherein theparticles are disposed within the fibers of the plurality of meshes inan amount up to about 30 weight percent, based on the total weight ofthe fibers plus the particles.
 7. The wound dressing of claim 1, whereinthe particles are disposed between the fibers of the mesh.
 8. The wounddressing of claim 1, wherein the biodegradable polymer fibers compriseone or more antimicrobial polymer fibers.
 9. The wound dressing of claim1, wherein the biodegradable polymer fibers comprise one or more ofchitosan, carboxymethyl chitosan (CMC), and poly(ethylene oxide). 10.The wound dressing of claim 1, wherein the biodegradable polymer fiberscomprise conductive polymer fibers comprising one or more ofpolypyrrole, polyaniline, or a poly-thiophene.
 11. The wound dressing ofclaim 1, wherein the first active agent comprises a growth factor. 12.The wound dressing of claim 1, wherein the biodegradable polymerparticles comprise one or more of a polyester, polylactic acid,polyglycolic acid, polylactic-co-glycolic acid, polycaprolactone, andcopolymers thereof.
 13. The wound dressing of claim 1, wherein thesecond active agent comprises a growth factor.
 14. The wound dressing ofclaim 1, wherein the plurality of meshes comprises a first mesh having ahydrophobicity and/or hydrophilicity different from a second mesh.
 15. Amethod of treating a wound comprising: applying the wound dressing ofclaim 1 to a surface of the wound.
 16. The method of claim 15 furthercomprising at least partially degrading the biodegradable polymer fibersof the wound dressing to release the first active agent into the wound.17. The method of claim 16 further comprising at least partiallydegrading the biodegradable polymer particles of the wound dressing torelease the second active agent into the wound, wherein the secondactive agent is released after the first active agent is released. 18.The method of claim 17, wherein the first active agent comprises agrowth factor for angiogenesis.
 19. The method of claim 18, wherein thesecond active agent comprises a growth factor for wound healing or bonegrowth.
 20. A wound dressing comprising: a plurality of meshes formedfrom a plurality of biodegradable polymer fibers; wherein thebiodegradable polymer fibers comprise a first active agent; wherein theplurality of meshes further comprise a plurality of biodegradablepolymer particles; wherein the biodegradable polymer particles comprisea second active agent; wherein an in vivo or in vitro release profile ofthe first active agent differs from an in vivo or in vitro releaseprofile of the second active agent; such that the in vivo or in vitrorelease profile of the first active agent and the in vivo or in vitrorelease profile of the second active agent overlap by less than about70%; wherein the biodegradable polymer fibers have an average diameterof between about 10 nm and about 500 nm and comprise one or morepolymers comprising a citrate moiety; wherein the biodegradable polymerparticles have an average size in one, two or three dimensions ofbetween about 10 nm and about 200 nm; wherein the plurality of meshesare arranged in a stack wherein the stack has a top surface and a bottomsurface, wherein the bottom surface is defined by a mesh placed on awound and the top surface is defined by a mesh disposed farthermost fromthe wound, wherein the stack having a property gradient in a z-directionof the stack such that the property increases from the bottom surface tothe top surface or such that the property decreases from the bottomsurface to the top surface of the stack; wherein the z-direction isdefined in a stacking direction; and wherein the wound dressing providesa fibrous scaffold for supporting cell growth; wherein the plurality ofmeshes is at least three meshes, and wherein the property is defined bya mesh porosity or a mesh perforation.
 21. The wound dressing of claim20, wherein the plurality of meshes comprises a first perforated meshand a second perforated mesh arranged in the stack, the first meshhaving a greater degree of perforation than the second mesh.
 22. Thewound dressing of claim 21, wherein the first perforated mesh has anaverage size perforation size between 10 μm to about 10 mm.
 23. Thewound dressing of claim 21, wherein the first perforated mesh has aperforation density between about 10 perforations/cm² to about 200perforations/cm².
 24. The wound dressing of claim 20, wherein theplurality of meshes comprises a first porous mesh and a second porousmesh arranged in the stack, the first mesh having a greater degree ofporosity than the second mesh, and wherein the first porous mesh has aporosity between about 10% to about 90%.
 25. The wound dressing of claim20, wherein the particles are disposed within the fibers of theplurality of meshes in an amount up to about 30 weight percent, based onthe total weight of the fibers plus the particles.
 26. The wounddressing of claim 20, wherein the particles are disposed between thefibers of the mesh.
 27. The wound dressing of claim 20, wherein thebiodegradable polymer fibers comprise one or more antimicrobial polymerfibers.
 28. The wound dressing of claim 20, wherein the biodegradablepolymer fibers comprise one or more of chitosan, carboxymethyl chitosan(CMC), and poly(ethylene oxide).
 29. The wound dressing of claim 20,wherein the biodegradable polymer fibers comprise conductive polymerfibers comprising one or more of polypyrrole, polyaniline, or apoly-thiophene.
 30. The wound dressing of claim 20, wherein the firstactive agent comprises a growth factor.
 31. The wound dressing of claim30, wherein the first active agent comprises a growth factor forangiogenesis.
 32. The wound dressing of claim 20, wherein thebiodegradable polymer particles comprise one or more of a polyester,polylactic acid, polyglycolic acid, polylactic-co-glycolic acid,polycaprolactone, and copolymers thereof.
 33. The wound dressing ofclaim 20, wherein the second active agent comprises a growth factor. 34.The wound dressing of claim 33, wherein the second active agentcomprises a growth factor for wound healing or bone growth.
 35. Thewound dressing of claim 20, wherein the plurality of meshes comprises afirst mesh having a hydrophobicity and/or hydrophilicity different froma second mesh.
 36. The wound dressing of claim 20, wherein the wounddressing exhibits a transfer of mechanical loads from the plurality ofmeshes to a regenerated biological tissue.
 37. A method of treating awound comprising: applying the wound dressing of claim 20 to a surfaceof the wound.